Invasive infections due to Aspergillus fumigatus are increasing, and despite new therapies are associated with greater than 50% mortality. Invasion of blood vessels is a hallmark pathologic feature of invasive aspergillosis and plays a critical role in the development of local and disseminated disease. This process of angioinvasion and subsequent dissemination is unique to A. fumigatus and a handful of other pathogenic molds, and its mechanism is not well understood. Histopathologic studies suggest that there are several key steps by which the organism gains entry to and subsequently exits from the vascular system. First, hyphae from an infectious focus must invade the abluminal surface of the blood vessel, and penetrate endothelial cells to gain access to the blood vessel lumen. Next, hyphal fragments are borne by the bloodstream to distal sites where they adhere to and penetrate the luminal surface of the endothelial cells lining the blood vessel and thereby invade the deep organs. Our overall hypothesis is that the interactions of A. fumigatus with the vascular endothelium are critical to the pathogenesis of invasive aspergillosis. We propose to study the mechanisms underlying angioinvasion and hematogenous dissemination by first developing an in vitro culture system that will enable us to study the interactions of A. fumigatus with both the abluminal and luminal surfaces of the endothelial cell. Using this system, we will investigate the mechanisms of endothelial cell invasion, stimulation, and injury when hyphae infect the abluminal vs. luminal surfaces of the endothelial cell. We will also characterize the parameters that govern the luminal adherence of A. fumigatus hyphae to endothelium and subendothelial matrix proteins. Finally, we will validate our in vitro results using two new murine models of invasive pulmonary and hematogenously disseminated aspergillosis. These studies are designed to lay the essential foundation for a comprehensive research program aimed at elucidating the molecular mechanisms by which A. fumigatus invades blood vessels and hematogenously disseminates.